Cryobiology最新文献

{"title":"Mitochondrial uncoupling and glycolysis stimulation are beneficial for kinematics, functionality and oxidative homeostasis of cryopreserved ram sperm","authors":"Álvaro de Miranda Alves ,&nbsp;João Diego de Agostini Losano ,&nbsp;Roberta Ferreira Leite ,&nbsp;Bruno Rogério Rui ,&nbsp;Daniel de Souza Ramos Angrimani ,&nbsp;Thais Rose dos Santos Hamilton ,&nbsp;Camilla Mota Mendes ,&nbsp;Mayra Elena Ortiz D'Avila Assumpção ,&nbsp;Marcilio Nichi","doi":"10.1016/j.cryobiol.2025.105236","DOIUrl":"10.1016/j.cryobiol.2025.105236","url":null,"abstract":"<div><div>The aim of the present study was to improve bioenergetics and oxidative status of cryopreserved ram sperm by uncoupling mitochondrial activity and stimulating glycolysis. To verify a potential synergism between mitochondrial uncoupling and glycolysis stimulation, as well as to determine the optimal concentrations of the respective treatments, the study was divided into two experiments. In Experiment 1, ejaculates from eight rams were diluted with the commercial extender (Botubov®), supplemented with the mitochondrial uncoupler CCCP (0, 1, 10, and 20 μM), with or without 5 mM glucose, and then subjected to cryopreservation. After thawing, sperm function and oxidative status analyses were conducted to determine the optimal CCCP concentration, which was selected for Experiment 2. In Experiment 2, ejaculates from seven rams were diluted with the commercial extender (Botubov®) and supplemented with CCCP at doses of 0, 2.5, 5, and 10 μM, with or without 5 mM glucose. After thawing, an analysis of sperm bioenergetics was performed. Differences between treatments were assessed using ANOVA, followed by LSD mean comparison test for the combination of factors. In both experiments, total and progressive motility were higher in the CCCP 10 μM + glucose 5 mM group. This same group exhibited less susceptibility to lipid peroxidation, lower DNA fragmentation (Experiment 1), and greater mitochondrial activity (Experiment 2). Furthermore, treatments with only CCCP were deleterious to sperm. In conclusion, the use of the mitochondrial uncoupler CCCP at a dose of 10 μM combined with 5 mM glucose was promising in improving post-thaw sperm attributes.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105236"},"PeriodicalIF":2.3,"publicationDate":"2025-03-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143684446","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of post-transfer fertility of vitrified goat morulae and blastocysts using an innovative protocol in micropipette tips","authors":"María Macarena Bruno-Galarraga ,&nbsp;Jimena Fernandez ,&nbsp;Agustí Noya ,&nbsp;Alejandro Gibbons ,&nbsp;Marcela Cueto","doi":"10.1016/j.cryobiol.2025.105234","DOIUrl":"10.1016/j.cryobiol.2025.105234","url":null,"abstract":"<div><div>In 2022, we developed an innovative protocol consisting of three vitrification solutions with the addition of sucrose to vitrify goat embryos in micropipette tips (S-VitriTip) and obtained promising <em>in vitro</em> results. However, to date, this protocol has not been tested <em>in vivo</em> in recipient females. The study aimed to evaluate the embryo survival and post-transfer fertility of goat morulae and blastocysts vitrified by the S-VitriTip protocol. Embryo recovery from donor goats (<em>n</em> = 23) was carried out on Days 8 and 9 after sponge removal. Immediately after recovery, morulae and blastocysts were vitrified using the S-VitriTip protocol. Briefly, all embryos were exposed to three different increasing solutions of cryoprotectants (glycerol, ethylene glycol, sucrose), loaded into micropipette tips and stored in liquid nitrogen. After warming, embryos were transferred in pairs into recipient goats (<em>n</em> = 47); morulae and blastocysts were transferred on the same day they were recovered, either Day 8 or Day 9 after sponge removal. On day 25 after embryo transfer, ultrasound diagnosis was performed. No significant differences were observed on embryo survival nor on post-transfer fertility according to embryo stage or day of embryo recovery (42 and 50 %, respectively; P &gt; 0.05). Embryo survival and post-transfer fertility of vitrified morulae of 30 % and 40 % were obtained, highlighting our study as one of the first to report efficient reproductive rates through the <em>in vivo</em> transfer of cryopreserved goat morulae, thus demonstrating the effectiveness of this innovative vitrification protocol.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105234"},"PeriodicalIF":2.3,"publicationDate":"2025-03-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143673561","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Bacterial survival below zero: Impact of storage time on bacterial viability in bull semen","authors":"Aleksandar Cojkic ,&nbsp;Ingrid Hansson ,&nbsp;Jane M. Morrell","doi":"10.1016/j.cryobiol.2025.105233","DOIUrl":"10.1016/j.cryobiol.2025.105233","url":null,"abstract":"<div><div>Although freezing methods have been optimized for preserving sperm integrity, their effectiveness in sustaining bacterial viability is unknown. Therefore, culturing thawed semen samples might not give an accurate picture of the bacteria in the original sample. The aim of this study was to assess how cryopreservation and storage duration influence bacterial populations and the survival of distinct bacterial species. Semen samples were collected from 14 bulls, samples were diluted in equal proportions of antibiotic-free semen extender and transported to the laboratory at 6 °C overnight. Aliquots of semen were cultured within 24 h after semen collection on Plate Count Agar to calculate number of bacteria, and blood agar plates (5 % bovine blood) for identification of bacterial species. The remaining samples were diluted 1:1 in Brain Heart Infusion (BHI) broth with 30 % glycerol and stored at −80 °C. The frozen samples were thawed and cultured for quantification of bacteria as described for fresh semen, after 6 and 13 days at −80 °C. The isolated bacteria were re-cultured on blood agar, incubated for one day at 37 °C before identification by Matrix assisted laser desorption ionization-time of flight mass spectrometry. Total bacterial counts remained consistent across fresh and cryopreserved samples regardless of storage duration. A total of 31 bacterial species were identified, with 20 detected in fresh samples, 16 present after 6 days of storage, and 18 observed after 13 days. Ten species persisted across all time points, while others were unique to a specific sampling day, including nine species on day 1, two species on day 6, and five species on day 13. These findings suggest that while cryopreservation does not alter the overall bacterial load, the survival of individual species varies depending on storage conditions.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105233"},"PeriodicalIF":2.3,"publicationDate":"2025-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143644327","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Vitrification of porcine intact femoral condyle through different protocol lengths and loading strategies","authors":"Mohammadhamed Shahsavari ,&nbsp;Korinne M. Tirones ,&nbsp;Marggi A. Patel ,&nbsp;Janet A.W. Elliott ,&nbsp;Nadr M. Jomha","doi":"10.1016/j.cryobiol.2025.105215","DOIUrl":"10.1016/j.cryobiol.2025.105215","url":null,"abstract":"<div><div>Cryopreservation of articular cartilage (AC) allografts is a potential candidate to solve the shortage of fresh osteochondral allografts to treat large AC defects. This technique requires optimization to address issues related to the actual process of cryopreservation and the translation of the technology to tissue banks. In the present study, we compared different vitrification protocols consisting of multiple cryoprotective agent (CPA) loading timings and loading strategies to achieve successful vitrification-rewarming protocols for porcine intact femoral condyles. Porcine medial femoral condyles (n = 6) were used to test 5 vitrification protocols differing in lengths of time and the method of CPA loading. The effects of these protocols on post-warming normalized chondrocyte viability, metabolic activity, and tissue cracking frequency were documented. We demonstrated that all 5 CPA loading-vitrification protocols resulted in high normalized chondrocyte viability (&gt;88 % of Fresh control) post vitrification in porcine intact femoral condyles. Moreover, the Room Temperature Gel (RG) protocol showed high viability and metabolic activity of chondrocytes post-warming, and also decreased cracking frequency in AC after rewarming of vitrified intact femoral condyles. To facilitate translation of this technology to tissue banks, we showed that by shortening (total time of 15 min in Protocol RG) or combining our established Protocol 8 (430 min) with a hydrogel, including use of cryobags in liquid nitrogen vapour as the conventional storage method in tissue banks, we could introduce two reproducible protocols for clinical applications involving vitrified osteochondral tissue.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105215"},"PeriodicalIF":2.3,"publicationDate":"2025-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143476328","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"BGP-15 improves quality of goat sperm by mitigating oxidative stress during cryopreservation","authors":"Fuqin Liu ,&nbsp;Jianjun Dai ,&nbsp;Jun Gao ,&nbsp;Mengqian He ,&nbsp;Jiehuan Xu ,&nbsp;Caifeng Wu ,&nbsp;Shushan Zhang ,&nbsp;Xing Zhu ,&nbsp;Lingwei Sun","doi":"10.1016/j.cryobiol.2025.105232","DOIUrl":"10.1016/j.cryobiol.2025.105232","url":null,"abstract":"<div><div>The objective of this study was to evaluate the effect of the addition of BGP-15 ((Z)-N'-(2-hydroxy-3-(piperidin-1-yl) propoxy) nicotinimidamide) on goat sperm during cryopreservation. The semen from six rams was diluted with a soybean lecithin-based extender containing different doses of BGP-15 (0, 50, 100, 150, and 200 μM). After the freeze-thawing process, sperm characteristics, plasma membrane integrity and functionality, acrosomal status, mitochondria activity, apoptosis status, and gene expression of post-thawed ram spermatozoa were assessed. Compared to fresh spermatozoa, the ultrastructure of frozen spermatozoa was damaged, and frozen spermatozoa had a reduced number of duplex microtubules with partially asymmetric distributions, localized disappearance of the central microtubule, and ambiguous submicrotubule structures. Moreover, the 100 μM BGP-15 treatment group had the highest percentage of viability, plasma membrane and acrosome integrities, and mitochondrial activity of frozen-thawed spermatozoa when compared to the control (P &lt; 0.05). In the Computer-Assisted Semen Analyzer, sperm motility parameters were significantly increased in the BGP-15 treatment group against the control group (P &lt; 0.05), except wobble. Similarly, BGP-15 treatment increased the levels of T-AOC, SOD, CAT, and GHS-Px and decreased the level of ROS which compared to controls (P &lt; 0.05). Only the 100 μM BGP-15 treatment group had a higher MDA level than the control group (P &lt; 0.05). In all BGP-15 treatment groups, the mRNA expressions of the <em>ROMO1</em> gene were significantly reduced compared to controls (P &lt; 0.05), and the <em>mRNA</em> expressions of the <em>SMCP, MnSOD</em>, <em>and CuZnSOD</em> genes were significantly increased (P &lt; 0.05). While the difference in the <em>mRNA</em> expression of the <em>SMOX</em> gene between the 50 μM BGP-15 treatment group and the control group was not significant (P &gt; 0.05). Moreover, the apoptosis rate of freeze-thawed spermatozoa significantly decreased in the 100 μM BGP-15 treatment group compared to the control (P &lt; 0.05), while the MMP of freeze-thawed spermatozoa significantly enhanced in the 100 μM BGP-15 treatment group (P &lt; 0.05). In conclusion, BGP-15 enhances cryo-protective effects on goat spermatozoa, and 100 μM BGP-15 addition to the extender during cryopreservation is beneficial to the goat breeding industry.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105232"},"PeriodicalIF":2.3,"publicationDate":"2025-03-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143620760","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Evaluation of the effect of human testicular cell conditioned media on the in vitro development of follicles from cryopreserved human ovarian cortical pieces. A potential approach for fertility preservation for cancer patients","authors":"Mohammad Ali Khalili ,&nbsp;Behrouz Aflatoonian ,&nbsp;Mohammad Reza Mirzaei ,&nbsp;Mahin Izadi ,&nbsp;Mojgan Noroozi Karimabad ,&nbsp;Fatemeh Asadi ,&nbsp;Mahboubeh Vatanparast","doi":"10.1016/j.cryobiol.2025.105218","DOIUrl":"10.1016/j.cryobiol.2025.105218","url":null,"abstract":"<div><div>If in vitro culture conditions could be improved, in vitro growth of cryopreserved ovarian tissue and isolated follicles could become an alternative to ovarian tissue transplantation. This study evaluated two cryopreservation methods, slow cooling and vitrification, and two in vitro culture methods, to determine their effect on the in vitro growth of human ovarian follicles. After warming, the OT pieces (1 × 10 × 3 mm) were cultured in either routine culture medium (DMEM), or DMEM supplemented 50:50 with media previously used for human testicular cell culture (hTCCM) to serve as a source of growth factors. Several parameters were evaluated during culture including follicle recruitment, growth, morphology, diameter, hormone production, and gene expressions (PTEN, BMP-15, PDGF, GDF-9, and GAPDH). The follicular morphology and hormonal secretion were comparable for both cryopreservation methods and both culture methods (P &gt; 0.05). Follicle recruitment was accelerated in the vitrified group, in the presence of hTCCM (P &gt; 0.05). After 7 days of culture, the follicle diameter was greater in the slow/hTCCM compared to the vitrification DMEM group. Similarly only one gene expression profile, BMP-15, in the slow/hTCCM differed significantly from another treatment group (Vit DMEM). At the end of culture (over 21 days), three immature, low-quality oocytes were the achievement. In conclusion, a very quick and easy vitrification protocol gave comparable results to the slow cooling method. We also established that the laboriously prepared hTCCM supplement did not benefit the outcomes. Further work is needed before in vitro maturation of cryopreserved ovarian cortical pieces can be used clinically.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105218"},"PeriodicalIF":2.3,"publicationDate":"2025-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143609661","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The effect of cryopreservation on enamel microcracks – A μCT analysis using a deep learning algorithm","authors":"Noëmi M.C. De Roo ,&nbsp;Pierre Kibleur ,&nbsp;Liesbeth Temmerman ,&nbsp;Ronald M.H. Verbeeck ,&nbsp;Matthieu N. Boone ,&nbsp;Guy A.M. De Pauw","doi":"10.1016/j.cryobiol.2025.105207","DOIUrl":"10.1016/j.cryobiol.2025.105207","url":null,"abstract":"<div><div>To date, the effect of cryopreservation on microcracks in the dental enamel remains unclear. These enamel microcracks are very thin, at the limit of visibility and their segmentation is beyond the capabilities of traditional image analysis. The objective of the present study was to investigate the effect of cryopreservation on enamel microcracks with a μCT analysis using a deep learning algorithm.</div><div>A manual annotation was performed to construct a high-quality training and testing dataset. A 4-phase semantic segmentation U-Net architecture neural network was trained in Dragonfly (ORS systems, Canada) and then applied on a sample of 5 teeth before and after cryopreservation, enabling for the first time a direct evaluation of the formation and evolution of enamel cracks caused by cryopreservation.</div><div>Qualitatively, the segmentation results were very satisfying and cracks as thin as 2–3 voxels wide could be segmented automatically. All teeth presented enamel microcracks, without propagation into the dentin. Similar crack patterns were observed in all teeth and may be related to the use of forceps during extraction. In the post-cryopreservation scans the damage extended, and new smaller cracks appeared on the occlusal surface of the tooth. Quantitatively, the average crack/enamel ratio was 0.066 ± 0.021 % before cryopreservation, and 0.087 ± 0.018 % after cryopreservation.</div><div>The present study presents the first scalable yet precise method to quantify clinically relevant tooth damage after cryopreservation. As such, this method also opens the way to future in-depth studies on dental enamel in many dental fields.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105207"},"PeriodicalIF":2.3,"publicationDate":"2025-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143390452","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Effect of long-term storage on the quality of cryopreserved sperm of the Pangasius nasutus (Bleeker, 1863)","authors":"Nurizzati Idris ,&nbsp;Donald Torsabo ,&nbsp;Muhammad Yazed Abduh ,&nbsp;Ambok Bolong Abol-Munafi ,&nbsp;Noordiyana Mat Noordin ,&nbsp;Ivan Chong Chu Koh","doi":"10.1016/j.cryobiol.2025.105219","DOIUrl":"10.1016/j.cryobiol.2025.105219","url":null,"abstract":"<div><div>In the present study, we investigated the effects of storage for 3, 6, 9 and 12 months on cryopreserved sperm of <em>P. nasutus</em>, in 10 % MeOH as a cryoprotectant with 90 % 0.9 % NaCl as an extender. Sperm quality on motility, fertilization, hatching, and deformity rates were investigated using 9-months cryopreserved sperm. Determination on the effects of different sperm to egg ratios also was evaluated using 1-year cryopreserved sperm. Post-thaw motility (PTM) of cryopreserved sperm within storage period was significantly lower compared to the initial motility of fresh sperm (51.67 ± 2.4 %). However, there was no significant change of PTM during 12 months of storage (3 months, 33.3 ± 2.33 %; 6 months, 32.0 ± 2.52; 9 months, 32.67 ± 1.76; 12 months, 33.33 ± 1.67). In addition, the motility duration was unaffected by storage as there was no significant difference compared to fresh samples. Besides that, there were no significant differences in motility, fertilization and deformity rates for fresh and 9-months cryopreserved sperm except for hatching rate. Samples stored for 1 year resulted in high fertilization across all sperm to egg ratios, showing no difference compared to fresh sperm. Cryopreserved sperm exhibited a significant higher hatching rate (9.24 ± 4.68 %) at the sperm to egg ratio of 300,000:1 compared to 150,000:1 (3.44 ± 1.7 %). However, no significant difference in deformity rates for fresh and cryopreserved at sperm to egg ratio of 300,000:1. The newly developed cryopreservation protocol using methanol successfully preserved sperm quality, maintaining fertilization rates comparable to fresh sperm. This highlights its potential application in sustainable aquaculture and conservation strategies for <em>Pangasius nasutus</em>.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105219"},"PeriodicalIF":2.3,"publicationDate":"2025-03-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143593992","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Potential application of acid-form sophorolipids in cell cryopreservation","authors":"Thi Nhu Trang Nguyen,&nbsp;Yuki Saito,&nbsp;Motoki Tatsumi,&nbsp;Masashi Yamamoto,&nbsp;Yoshihiko Hirata","doi":"10.1016/j.cryobiol.2025.105228","DOIUrl":"10.1016/j.cryobiol.2025.105228","url":null,"abstract":"<div><div>In cryopreservation, which is an important process for the long-term storage and transport of cells, cryopreservation solutions containing cryoprotectants are usually used to protect cells from damage. However, most cryoprotectants have non-negligible cytotoxicity and side effects that greatly limit their applications in clinical use. This has led to an increased need for more biocompatible cryoprotectants, and interest in bioinspired cryoprotectants is increasing. In this study, we have discovered the potential of using natural acid-form sophorolipids (aSL) as a cryoprotectant, which suppresses ice formation and reduces osmotic stress. The solution containing aSL showed the smallest size of ice crystals compared to the solutions containing other cryoprotectants, such as dimethyl sulfoxide (Me2SO), glycerol (GL), ethylene glycol (EG), and propylene glycol (PG). By adding aSL to a hypertonic culture medium, cell viability was significantly improved. This finding suggests an opportunity to develop low-toxicity and efficient reagents for cell cryopreservation in the future.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105228"},"PeriodicalIF":2.3,"publicationDate":"2025-03-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143593993","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Effect of vitrification on in vitro developmental competence of rat testicular tissue","authors":"Mina Sharbatoghli ,&nbsp;Samira Hajiaghalou ,&nbsp;Nafiseh Sadat Deheshkar Gooneh Farahani ,&nbsp;Samaneh Aghajanpour ,&nbsp;Abdolhossein Shahverdi ,&nbsp;Mojtaba Rezazadeh Valojerdi ,&nbsp;Bita Ebrahimi","doi":"10.1016/j.cryobiol.2025.105213","DOIUrl":"10.1016/j.cryobiol.2025.105213","url":null,"abstract":"<div><div>Cryopreservation of testicular tissue has been proposed as a potential technique for preserving fertility in pre-pubertal boys with various malignancies. The present study aimed to compare the effects of two vitrification techniques—solid surface vitrification (SSV) and needle-immersed vitrification (NIV)—on the integrity, development, cell viability, and apoptosis of rat testicular tissue. Testes from 4-week-old Wistar rats underwent a two-step vitrification process. Tissue pieces were allocated to either the SSV or NIV group. Equilibration involved a solution containing 7.5 % dimethyl sulfoxide (DMSO) and 7.5 % ethylene glycol (EG), followed by a vitrification solution with 0.07 mol/L sucrose, 15 % DMSO, and 15 % EG. The optimal protocol was determined after vitrification using either the NIV or SSV technique. Samples from the control and selected vitrification (SSV) groups were cultured for 3 weeks. Tissue integrity, cell viability, apoptosis, and gene expression were evaluated using hematoxylin and eosin staining, trypan blue staining, annexin V-PI staining, and real-time PCR. Morphological changes were more pronounced in the NIV group compared to the SSV group (P &lt; 0.05). Although the percentage of viable cells did not significantly differ between the NIV and SSV groups, it was slightly higher in the SSV group. Thus, SSV was identified as the optimum vitrification method. Real-time PCR analysis revealed altered gene expression: spermatogonial-related genes (<em>Lrp4</em>, <em>Egr3</em>, <em>Nanos</em>, <em>Gfra1</em>, <em>C-kit</em>, and <em>Sohlh1</em>) were significantly decreased in the SSV group, while somatic-cells-related genes (<em>Gdnf</em>, <em>Csf1</em>, and <em>Fgf2</em>) were higher. Overall, SSV appears suitable for rat testis tissue vitrification, although it induces some molecular changes. Optimization of the culture medium is essential to support successful spermatogenesis.</div></div>","PeriodicalId":10897,"journal":{"name":"Cryobiology","volume":"119 ","pages":"Article 105213"},"PeriodicalIF":2.3,"publicationDate":"2025-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143425143","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}